ALDH Expression in PCa Cell lines

C4 C4-2 C4-2B PC-3 PC-3M- PC-3M- DU145 Pro4luc Pro4lucBIII ALDH1A1 - - +++ ++ - ++ ++ ALDH2 +++ +++ +++ - - - + ALDH3A1 - + - + - - + ALDH3A2 ++++ ++++ ++++ +++ +++ +++ +++ ALDH4A1 +++ +++ +++ + + + ++ ALDH5A1 ++ ++ +++ + + + + ALDH6A1 ++++ +++ +++ + + + + ALDH7A1 + + ++ +++ ++ +++ +++ ALDH8A1 + ------ALDH9A1 +++ +++ +++ ++ ++ +++ ++ ALDH16A1 + + ++ + - + - ALDH18A1 +++ ++++ ++++ ++ +++ +++ +++

Supplementary Table I. Expression levels of ALDH isoforms in prostate cancer cell lines. Expression levels of the ALDH isoforms were assessed with real time qPCR in prostate cancer cell lines Data are classified in levels according to their relative transcript abundance (RTA: 10000/2^ΔCt). RTA <1 = - (Ct value ~38); RTA 1-20 = + (Ct value ~34); RTA 21-100 = ++ (Ct value ~30); RTA 101-500 = +++ (Ct value ~26); RTA > 500 =++++ ( housekeeping gene level Ct value ~22). ND = not determined, not-significant amounts of material.

ALDH Expression in Prostate Cancer

PCa primary tumor PCa primary cultures #44 #99 #62#69 123 583 697 233 549 069B 567B ALDH1A1 ++ + ND ND ++++ ND ND ND ND ND ND ALDH2 + ++ ND ND ++++ ND ND ND ND ND ND ALDH3A1 ++++ ++++ +++ ++ ++ +++ ++++ ++++ +++ ++ + ALDH3A2 +++ +++ +++ ++++ + ++++ ++++ ++++ ++++ ++++ ++++ ALDH4A1 ++ +++ ++ ++ +++ ND ND ND ND ND ND ALDH5A1 ++ +++ ++ ++ - ND ND ND ND ND ND ALDH6A1 ++ + ND ND ++++ ND ND ND ND ND ND ALDH7A1 ++++ ++++ +++ +++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ALDH8A1 + + ND ND + ND ND ND ND ND ND ALDH9A1 ++ ++ ND ND ++++ ND ND ND ND ND ND ALDH16A1 +++ ++ ND ND ++++ ND ND ND ND ND ND ALDH18A1 - - +++ +++ - ++++ ++++ ++++ ++++ +++++++

Supplementary Table II. Expression levels of ALDH isoforms in primary prostate cancer tissue samples and primary cultures. Expression levels of the ALDH isoforms were assessed with real time qPCR in primary prostate cancer tissue samples (left panel) and primary cultures (right panel). Data are classified in levels according to their relative transcript abundance. RTA <1 = - (Ct value ~38); RTA 1-20 = + (Ct value ~34); RTA 21-100 = ++ (Ct value ~30); RTA 101-500 = +++ (Ct value ~26); RTA > 500 =++++ ( housekeeping gene level Ct value ~22). ND = not determined, not-significant amounts of material.

Supplemental Table III. IHC antibodies

Primary antibody Antigen retrieval Dilution Supplier Mouse monoclonal anti- Not necessary 1:100 BD Biosciences human ALDH1A1 Rabbit polyclonal anti- Heating samples in 0.1 M citrate 1:50 Sigma ATLAS human ALDH3A2 buffer, pH6, in a microwave oven for 10 min Rabbit monoclonal anti- Not necessary 1:100 AbCaM human ALDH7A1 Rabbit polyclonal anti- Heating samples in 0.1 M citrate 1:50 Sigma ATLAS human ALDH18A1 buffer, pH6, in a microwave oven for 10 min Supplementary Table III. ALDH antibodies used for Immunohistochemical analysis with the necessary antigen retrieval methods, required dilution and suppliers.

Table IV. Real time-qPCR primers

Gene 5’ Sequence 3’ Sequence ALDH1A1 TGGCTTATCAGCAGGAGTGT GCAATTCACCCACACTGTTC ALDH2 CGAGGTCTTCTGCAACCAG GCCTTGTCCACATCTTCCTT ALDH3A1 CAGGACGAGCTCTACATCCA GGCTTGAGGACCACTGAGTT ALDH3A2 CCCCTTCGTTCTCACCATTC CTGAGGGAGAAGCTTTGCCA ALDH4A1 GGACGTGCAGTACCAAGTGT TAGGCTTCAGGTCCCACTCT ALDH5A1 AAGGATGACCTTGCCAGAAT GGCTTCCTCAGAGAACCACT ALDH6A1 CTGCAGGTTTCTTCCAAGGT ATTTCCCACCAATGAAGAGC ALDH7A1 CAACGAGCCAATAGCAAGAG GCATCGCCAATCTGTCTTAC ALDH8A1 GCAGAGTGCCAAATAGTGGA AACTCCTCCAGGGACTGCT ALDH9A1 GCTGCCAGGATAATAAGGGA GAAATGTCAATGTCCAAGCG ALDH16A1 AGGATCCCATCACAGGAGAG ACTCCAGCCCTTAAATGCC ALDH18A1 ATCAGTGCTGCAGAATCAGG TGGAATTGCCATTTCTTTCA TMPRSS:ERG CAGGAGGCGGAGGCGGA GGCGTTGTAGCTGGGGGTGAG CD44 TGGCACCCGCTATGTCCAG GTAGCAGGGATTCTGTCTG α2 integrin TTTGGTAGTGTGCTGTGTTC GACTCTTCCTTCCTCTTTCTTTAG CD133 AGAGCTTGCACCAACAAAGTACAC AAGCACAGAGGGTCATTGAGAGA GAPDH GACAGTCAGCCGCATCTTC GCAACAATATCCACTTTACCAGAG Supplementary Table IV. Exon-spanning real time-PCR primers designed with Primer Express software (Applied Biosystems, Rotkreuz, Switzerland)